At the time reactions such as amplification of nucleic acids (DNA, RNA, and the like) and the fragments thereof (oligonucleotides, nucleotides, and the like) are performed, in tests that require quantitativeness, such as the analysis of gene expression levels, it becomes necessary to perform the amplification such that the ratio of the relative amounts of the respective nucleic acids can be known. Consequently, by using the real-time PCR method, and by using an apparatus provided with a thermal cycler and a fluorescence spectrophotometer, analysis by electrophoresis is made unnecessary as a result of the generation process of the DNA amplification products in PCR being detected and analyzed in real time. Furthermore, as a DNA amplification method that performs amplification while maintaining the quantitativeness with respect to the ratio of the relative amounts of the DNA or RNA contained in the sample before amplification, the SPIA (Single Primer Isothermal Amplification) method is used. In the SPIA method, the linear DNA amplification method resulting from an isothermal reaction utilizing DNA/RNA chimera primer, DNA polymerase, and RNaseH has become used.
In a case where processing such as nucleic acid amplification, and measurements thereof are performed, conventionally, the target nucleic acid is separated and extracted from the sample by using a filter by means of a manual method, by using magnetic particles and adsorption on an inner wall of a vessel or a pipette tip by means of a magnetic field, or by using a centrifuge. The separated and extracted target compound is transferred and introduced into a reaction vessel together with a reaction solution by a manual method and the like, and upon sealing of the reaction vessel using a manual method and the like, at the time reactions are performed using a temperature control device for reactions, optical measurements are performed with respect to the reaction vessel using a light measuring device (Patent Document 1).
In a case where the processing is executed by a manual method, a large burden is forced on the user. Furthermore, in a case where the processing is executed by combining a dispenser, a centrifuge, a magnetic force device, a temperature controller, a device for sealing the reaction vessel, a light measurement device, and the like, there is a concern of the scale of the utilized devices increasing and of the work area expanding. In particular, in a case where a plurality of samples is handled, since it becomes necessary to separate and extract a plurality of target nucleic acids and for amplification to be to respectively performed, the labor thereof becomes even greater, and furthermore, there is a concern of the work area also expanding further.
Specifically, in a case where reactions of the nucleic acids (DNA, RNA, and the like) to be amplified, and the like, are performed within a plurality of reaction vessels and these reactions are monitored by optical measurements, the measurements are performed by successively moving a single measuring device to the respective reaction vessels by a manual method, or the measurements are performed by providing a measuring device to each of the respective reaction vessels beforehand.
In the former case where a single measuring device is used, when the measuring device is attempted to be manually moved to the apertures of the reaction vessels, there is a concern of subtle differences occurring in the measurement conditions for each reaction vessel as a result of subtle displacements or relative motions between the reaction vessel and the measuring device.
In the latter case where a measuring device is provided to each of the respective reaction vessels, although the positioning accuracy becomes high, there is a concern of the devices scale expanding, and of the manufacturing costs increasing. Furthermore, although it is preferable to seal the apertures of the reaction vessels at the time of temperature control and the measurements, it is time-consuming to perform sealing, or opening and closing, with respect to a plurality of reaction vessels by a manual method with a lid, and in particular, there is a concern of the lid becoming adhered to the vessel apertures such that it becomes difficult to easily open the lid, and of contamination occurring from the liquid attached to the inside of the lid dripping or splashing. Furthermore, there is a concern of providing a dedicated opening and closing mechanism of the lid, complicating the apparatus, and increasing the manufacturing costs (Patent Document 2).
As a device that automatically performs measurements without providing a measurement device for each of the respective reaction vessels, there is an device that, at the time temperature control of a microplate having a plurality of wells is performed by a thermal cycler, successive light measurements of the respective wells is performed by moving a detection module over the microplate (Patent Documents 3 and 4).
In this device, since the detection module itself is moved in a state in which it is supported by the thermal cycler, a load that accompanies the acceleration from the movement is imparted on the detection module, which has precision optical system elements and electronic circuits such as a photomultiplier tube, thus becoming the cause of noise or breakdowns of the measurement device, and furthermore, there is a concern of the device lifetime being shortened.
Moreover, since the detection module is supported by the microplate, or is supported by a lid body sealing the respective wells of the microplate and only moves in a horizontal direction, a fixed spacing between the respective wells and the measuring end of the measurement device is necessary. Therefore, since attenuation from the scattering of light, and the leakage or entry of light with respect to the adjacent wells cannot be completely blocked and prevented, there is a concern of a measurement with a high accuracy not being able to be performed.
Furthermore, since the detection module divides the light path using a half mirror at the time light from the vessels is received or is irradiated, there is a need to take a long light path length within the measurement device, and it has a problem in that there is a concern of the device scale becoming large.
In a case where the detection module is one that moves such that it passes the respective wells that are arranged on the microplate and the number of wells becomes large, there is a concern of the processing time becoming long due to the movement distance being long, and in addition, there is also a concern of the problems of the measurement device mentioned above occurring.
Moreover, at the time an optical measurement is performed on a sealed reaction vessel, there is a concern of the lid which has transparency, or the optical system elements, becoming cloudy from condensation, and the measurements becoming difficult.
Consequently, in order to perform nucleic acid amplification and the like, as a precondition thereof, specialized researchers or technicians become necessary, and this situation is preventing the generalization of genetic analysis and the expansion of clinical applications in hospitals, and the like.
Therefore, at the time of clinical use and the like, in order to prevent cross-contamination and to reduce user labor, and to easily perform the genetic analysis of nucleic acids from the extraction, the amplification, and further, by means of a measurement, then consistently automating the steps from extraction of the target compound, reacting such as amplifying, up to measurement, and miniaturization of the device, and the provision of an inexpensive, high-accuracy device are important.